Compounds

ABSTRACT

There are provided novel compounds of formula (I) ##STR1## wherein R 1  represents a 2-thienyl or 3-thienyl ring and R 2  represents C 1 to 4 alkyl and optical isomers and racemates thereof and pharmaceutically acceptable salts thereof; together with processes for their preparation, compositions containing them and their use in therapy. The compounds are selective inhibitors of the neuronal isoform of nitric oxide synthase.

This application is a 371 of PCT/SE98/00792 filed May 5, 1997.

FIELD OF THE INVENTION

This invention relates to new amidine derivatives, processes for theirpreparation, compositions containing them and their use in therapy.

BACKGROUND OF THE INVENTION

Nitric oxide is produced in mammalian cells from L-arginine by theaction of specific nitric oxide synthases (NOSs). These enzymes fallinto two distinct classes--constitutive NOS (cNOS) and inducible NOS(iNOS). At the present time, two constitutive NOSs and one inducible NOShave been identified. Of the constitutive NOSs, an endothelial enzyme(ecNOS) is involved with smooth muscle relaxation and the regulation ofblood pressure and blood flow, whereas the neuronal enzyme (ncNOS)serves as a neurotransmitter and appears to be involved in theregulation of various biological functions such as cerebral ischaemia.Inducible NOS has been implicated in the pathogenesis of inflammatorydiseases. Specific regulation of these enzymes should therefore offerconsiderable potential in the treatment of a wide variety of diseasestates.

Compounds of various structures have been described as inhibitors of NOSand their use in therapy has been claimed. See, for example, WO 95/09619(The Wellcome Foundation) and WO 95/11231 (G. D. Searle). The applicanthas previously disclosed in WO 95/05363 and WO 96/01817 amidinederivatives which are NOS inhibitors which display some selectivity forinhibition of the neuronal enzyme, ncNOS.

We now disclose a group of amidines that are within the generic scope ofWO 96/01817, but which are not specifically exemplified in WO 96/01817.These compounds display surprisingly advantageous properties and are thesubject of the present application.

DISCLOSURE OF THE INVENTION

According to the invention we provide a compound of formula (I) ##STR2##wherein: R¹ represents a 2-thienyl or 3-thienyl ring;

and R represents C 1 to 4 alkyl;

and optical isomers and racemates thereof and pharmaceuticallyacceptable salts thereof.

Preferably R¹ represents 2-thienyl.

Particularly preferred compounds of the invention include:

N-(2-methyl-1,2,3,4-tetrahydroisoquinolin-7-yl)-2-thiophenecarboximidamide;

N-(2-isopropyl-1,2,3,4-tetrahydroisoquinolin-7-yl)-2-thiophenecarboximidamide;

N-(2-ethyl-1,2,3,4-tetrahydroisoquinolin-7-yl)-2-thiophenecarboximidamide;

N-(2-propyl-1,2,3,4-tetrahydroisoquinolin-7-yl)-2-thiophenecarboximidamide;

N-(2-methyl-1,2,3,4-tetrahydroisoquinolin-7-yl)-3-thiophenecarboximidamide;

and pharmaceutically acceptable salts thereof.

A more especially preferred compound of the invention is:

N-(2-methyl-1,2,3,4-tetrahydroisoquinolin-7-yl)-2-thiophenecarboximidamide;

and pharmaceutically acceptable salts thereof.

Unless otherwise indicated, the term "C 1 to 4 alkyl" referred to hereindenotes a straight or branched chain alkyl group having from 1 to 4carbon atoms. Examples of such groups include methyl, ethyl, n-propyl,i-propyl, n-butyl, i-butyl and t-butyl.

The present invention includes compounds of formula (I) in the form ofsalts, in particular acid addition salts. Suitable salts include thoseformed with both organic and inorganic acids. Such acid addition saltswill normally be pharmaceutically acceptable although salts ofnon-pharmaceutically acceptable acids may be of utility in thepreparation and purification of the compound in question. Thus,preferred salts include those formed from hydrochloric, hydrobromic,sulphuric, phosphoric, citric, tartaric, lactic, pyruvic, acetic,succinic, fumaric, maleic, methanesulphonic and benzenesulphonic acids.

According to the invention, we further provide a process for thepreparation of compounds of formula (I), and optical isomers andracemates thereof and pharmaceutically acceptable salts thereof, whichcomprises:

(a) preparing a compound of formula (I) by reacting a correspondingcompound of formula (II) ##STR3## wherein R² is as defined above, with acompound of formula (III) or an acid addition salt thereof ##STR4##wherein R¹ is as defined above and L is a leaving group; (b) preparing acompound of formula (I) by reacting a corresponding compound of formula(IV) ##STR5## wherein R² is as defined above and HA is an acid, with acompound of formula (V) ##STR6## wherein R¹ is as defined above; (c)preparing a compound of formula (I) by reacting a compound of formula(VI) ##STR7## wherein R¹ is as defined above, with a compound of formula(VII)

    R.sup.2 --L                                                (VII)

wherein R² represents C 1 to 4 alkyl and L is a leaving group; or

(d) preparing a compound of formula (I) in which R² represents methyl byreacting a compound of formula (VI) with formaldehyde and formic acid;

and where desired or necessary converting the resultant compound offormula (I), or another salt thereof, into a pharmaceutically acceptablesalt thereof, or vice versa, and where desired converting the resultantcompound of formula (I) into an optical isomer thereof.

In process (a), the reaction will take place on stirring a mixture ofthe reactants in a suitable solvent, for example,N-methyl-2-pyrrolidinone or a lower alkanol such as ethanol, isopropanolor tertiary butanol, at a temperature between room temperature and thereflux temperature of the solvent. The reaction time will depend interalia on the solvent and the nature of the leaving group, and may be upto 48 hours; however it will typically be from 1 to 24 hours. Suitableleaving groups that L may represent include thioalkyl, sulphonyl,trifluoromethyl sulphonyl, halide, alkyl alcohols, aryl alcohols andtosyl groups; others are recited in `Advanced Organic Chemistry`, J.March (1985) 3rd Edition, on page 315 and are well known in the art.

In process (b), the reaction is preferably performed by refluxing amixture of the two compounds for several hours in the presence of asuitable solvent whereby the reaction temperature is high enough so thatcondensation takes place readily, but not sufficiently high to decomposethe amidine formed. The reaction temperature can vary from roomtemperature to about 250° C., although it is preferable to perform thereaction at temperatures from about 100° C. to 200° C. We find thato-dichlorobenzene is a particularly suitable solvent. We also find thatit is often useful to add 4-dimethylaminopyridine as a catalyst. Oncooling, two layers form, the solvent may be decanted, and the reactionworked up by addition of aqueous base. Alternatively, where thereactants are soluble in the solvent, the solvent may be evaporated offunder vacuum and the reaction mixture worked up by addition of water.The acid HA may be an organic or inorganic acid, for instance,hydrochloric, hydrobromic, hydroiodic, sulphuric, nitric, phosphoric,acetic, lactic, succinic, fumaric, malic, maleic, tartaric, citric,benzoic or methanesulphonic acid. We prefer that HA is a hydrohalicacid.

In process (c) the reaction will take place under standard conditions,for example by reacting the two compounds in an inert solvent such asDMF under basic conditions at a suitable temperature, typically roomtemperature, for a period of up to 72 hours or until the reaction iscomplete. We have frequently found it desirable to treat the amine withNaH before reacting with the compound of formula (VII). Suitable leavinggroups L are mentioned above. We prefer that L represents halide,particularly bromide.

In process (d), the reaction will typically take place on refluxing thereaction mixture for up to 4 hours or until reaction is complete.

Salts of compounds of formula (I) may be formed by reacting the freebase or a salt, enantiomer, tautomer or protected derivative thereof,with one or more equivalents of the appropriate acid. The reaction maybe carried out in a solvent or medium in which the salt is insoluble, orin a solvent in which the salt is soluble followed by subsequent removalof the solvent in vacuo or by freeze drying. Suitable solvents include,for example, water, dioxan, ethanol, isopropanol, tetrahydrofuran ordiethyl ether, or mixtures thereof. The reaction may be a metatheticalprocess or it may be carried out on an ion exchange resin.

The compounds of formula (II) may be prepared by reduction of acorresponding compound of formula (VIII) ##STR8## wherein R² is asdefined above.

The reduction reaction may be performed under a number of conditions,for example those described in J. March "Advanced Organic Chemistry" onpages 1103-1104. These include catalytic hydrogenation, use of Zn, Sn orFe metal, AlH₃ --AlCl₃, sulphides and others. We prefer to perform thereaction by hydrogenation at atmospheric pressure in the presence of apalladium and carbon catalyst until the reaction is complete, typically3 to 6 hours, or by reduction using zinc metal in acetic acid andmethanol.

Compounds of formula (VIII) may be prepared by nitration of a compoundof formula (IX) ##STR9## wherein R² is as defined above.

The nitration reaction will take place under conditions well known to aperson skilled in the art, for example, on treatment with nitric acidand sulphuric acid or potassium nitrate and sulphuric acid, optionallyin an inert organic solvent.

It may also be convenient to prepare compounds of formula (VIII) bynitration of a carbonyl or dicarbonyl derivative of a compound offormula (IX); which nitrated carbonyl or dicarbonyl derivative may bereduced to the desired compound of formula (VIII) using, for example,diborane.

Compounds of formula (VIII) and (IX), as well as certain carbonyl anddicarbonyl derivatives of compounds of formula (IX) just mentioned mayalso be prepared by one of the numerous methods for preparation ofbicyclic heterocyclic compounds.

Thus a compound of formula (X) ##STR10## may be prepared by ringexpansion of a cyclic ketone (XI) ##STR11## by treatment with sodiumazide in acid (Grunewald and Dahanukar, J. Heterocyclic Chem., 1994,31,1609-1617).

It will be apparent to a person skilled in the art that the compounds offormula (X) may also desirably be prepared in nitrated form. Nitrationmay be achieved by treatment of the non-nitrated analogue with nitricacid and sulphuric acid or potassium nitrate and sulphuric acid understandard conditions.

Intermediate compounds may be prepared as such or in protected form. Inparticular amine groups may be protected. Suitable protecting groups aredescribed in the standard text "Protective Groups in Organic Synthesis",2nd Edition (1991) by Greene and Wuts. Amine-protecting groups which maybe mentioned include alkyloxycarbonyl such as t-butyloxycarbonyl,phenylalkyloxycarbonyl such as benzyloxycarbonyl, or trifluoroacetate.Deprotection will normally take place on treatment with aqueous base oraqueous acid.

Compounds of formula (VIII) and (IX) in which R² represents C 1 to 4alkyl may also be prepared by alkylation of the corresponding N--Hcompound following process (c) above.

Compounds of formula (IV) may be prepared by analogous processes tothose described for the preparation of compounds of formula (II).Compounds of formula (IV) may be converted into corresponding compoundsof formula (II) by treatment with a base. Compounds of formula (II) maybe converted into corresponding compounds of formula (IV) by treatmentwith a protic acid HA, for example, one of those listed above.

Compounds of formula (III) are either known or may be prepared by knownmethods. For example, compounds of formula (III) in which L representsthioalkyl may be prepared by treatment of the corresponding thioamide offormula (XII) ##STR12## wherein R¹ is as defined above, with analkylhalide under conditions well known to a person skilled in the art.

Alternatively, the acid addition salts of compounds of formula (III)wherein L is thioalkyl may be prepared by reaction of a nitrile offormula (V) with an alkyl thiol and acid, for example hydrochloric acid,in a solvent such as dichloromethane or diethyl ether.

Compounds of formula (V), (VII), (X), (XI) and (XII) are either known ormay be prepared by conventional methods known per se.

It will be apparent to a person skilled in the art that it may bedesirable to protect an amine or other reactive group in an intermediatecompound using a protecting group as described in the standard text"Protective Groups in Organic Synthesis", 2nd Edition (1991) by Greeneand Wuts. Suitable amine-protecting groups are mentioned above.

The compounds of the invention and intermediates may be isolated fromtheir reaction mixtures, and if necessary further purified, by usingstandard techniques.

The compounds of formula (I) may exist in tautomeric, enantiomeric ordiastereoisomeric forms, all of which are included within the scope ofthe invention. The various optical isomers may be isolated by separationof a racemic mixture of the compounds using conventional techniques, forexample, fractional crystallisation or HPLC. Alternatively, theindividual enantiomers may be made by reaction of the appropriateoptically active starting materials under reaction conditions which willnot cause racemisation.

Intermediate compounds may also exist in enantiomeric forms and may beused as purified enantiomers, diastereomers, racemates or mixtures.

The compounds of general formula (I) possess useful nitric oxidesynthase inhibiting activity, and in particular, they exhibit goodselectivity for inhibition of the neuronal isoform of nitric oxidesynthase. They are thus useful in the treatment or prophylaxis of humandiseases or conditions in which the synthesis or oversynthesis of nitricoxide by nitric oxide synthase forms a contributory part. Examples ofsuch diseases or conditions include hypoxia, such as in cases of cardiacarrest, stroke and neonatal hypoxia, neurodegenerative conditionsincluding nerve degeneration and/or nerve necrosis in disorders such asischaemia, hypoxia, hypoglycemia, epilepsy, and in external wounds (suchas spinal cord and head injury), hyperbaric oxygen convulsions andtoxicity, dementia, for example, pre-senile dementia, Alzheimer'sdisease and AIDS-related dementia. Sydenham's chorea, Parkinson'sdisease, Huntington's disease, Amyotrophic Lateral Sclerosis,Korsakoff's disease, imbecility relating to a cerebral vessel disorder,sleeping disorders, schizophrenia, anxiety, depression, seasonalaffective disorder, jet-lag, depression or other symptoms associatedwith Premenstrual Syndrome (PMS), anxiety and septic shock. Thecompounds of formula (I) are also useful in the treatment andalleviation of acute or persistent inflammatory or neuropathic pain, orpain of central origin, and in the treatment or prophylaxis ofinflammation. Compounds of formula (I) are also predicted to showactivity in the prevention and reversal of tolerance to opiates anddiazepines, treatment of drug addiction and treatment of migraine andother vascular headaches. The compounds of the present invention mayalso show useful immunosuppressive activity, and be useful in thetreatment of gastrointestinal motility disorders, and in the inductionof labour. The compounds may also be useful in the treatment of cancersthat express nitric oxide synthase.

Compounds of formula (I) are predicted to be particularly useful in thetreatment or prophylaxis of hypoxia or stroke or ischaemia orneurodegenerative conditions or schizophrenia or of migraine or for theprevention and reversal of tolerance to opiates and diazepines or forthe treatment of drug addiction or for the treatment of pain andespecially in the treatment or prophylaxis of hypoxia or stroke orischaemia or neurodegenerative disorders or schizophrenia or pain. Weare particularly interested in conditions selected from the groupconsisting of hypoxia, ischaemia, stroke, pain, schizophrenia,Parkinson's disease, Huntington's disease and Amyotrophic LateralSclerosis.

For the treatment of Parkinson's disease, the compounds of formula (I)are expected to be particularly useful either alone, or in combinationwith other agents such as L-Dopa.

For the treatment of pain, the compounds of formula (I) are expected tobe particularly useful either alone, or in combination with other agentssuch as opiates, particularly morphine.

Prophylaxis is expected to be particularly relevant to the treatment ofpersons who have suffered a previous episode of, or are otherwiseconsidered to be at increased risk of, the disease or condition inquestion. Persons at risk of developing a particular disease orcondition generally include those having a family history of the diseaseor condition, or those who have been identified by genetic testing orscreening to be particularly susceptible to developing the disease orcondition.

Thus according to a further aspect of the invention we provide acompound of formula (I), or an optical isomer or racemate thereof or apharmaceutically acceptable salt thereof, for use as a medicament.

According to another feature of the invention we provide the use of acompound of formula (I) or an optical isomer or racemate thereof or apharmaceutically acceptable salt thereof, in the manufacture of amedicament for the treatment or prophylaxis of the aforementioneddiseases or conditions; and a method of treatment or prophylaxis of oneof the aforementioned diseases or conditions which comprisesadministering a therapeutically effective amount of a compound offormula (I), or an optical isomer or racemate thereof or apharmaceutically acceptable salt thereof, to a person suffering from orsusceptible to such a disease or condition.

For the above mentioned therapeutic indications, the dosage administeredwill, of course, vary with the compound employed, the mode ofadministration and the treatment desired. However, in general,satisfactory results are obtained when the compounds are administered toa human at a daily dosage of between 0.5 mg and 2000 mg (measured as theactive ingredient) per day, particularly at a daily dosage of between 2mg and 500 mg.

The compounds of formula (I), and optical isomers and racemates thereofand pharmaceutically acceptable salts thereof, may be used on their own,or in the form of appropriate medicinal formulations. Administration maybe by, but is not limited to, enteral (including oral, sublingual orrectal), intranasal, or topical or other parenteral routes. Conventionalprocedures for the selection and preparation of suitable pharmaceuticalformulations are described in, for example, "Pharmaceuticals--TheScience of Dosage Form Designs", M. E. Aulton, Churchill Livingstone,1988.

According to the invention, there is provided a pharmaceuticalformulation comprising preferably less than 95% by weight and morepreferably less than 50% by weight of a compound of formula (I), or anoptical isomer or racemate thereof or a pharmaceutically acceptable saltthereof, in admixture with a pharmaceutically acceptable diluent orcarrier. The formulation may optionally also contain a secondpharmacologically active ingredient such as L-Dopa, or an opiateanalgesic such as morphine.

We also provide a method of preparation of such a pharmaceuticalformulation which comprises mixing the ingredients.

Examples of such diluents and carriers are: for tablets and dragees:lactose, starch, talc, stearic acid; for capsules: tartaric acid orlactose; for injectable solutions: water, alcohols, glycerin, vegetableoils; for suppositories: natural or hardened oils or waxes.

Compositions in a form suitable for oral, that is oesophageal,administration include: tablets, capsules and dragees; sustained releasecompositions include those in which the active ingredient is bound to anion exchange resin which is optionally coated with a diffusion barrierto modify the release properties of the resin.

The enzyme nitric oxide synthase has a number of isoforms and compoundsof formula (I), and optical isomers and racemates thereof andpharmaceutically acceptable salts thereof, may be screened for nitricoxide synthase inhibiting activity by following procedures based onthose of Bredt and Snyder in Proc. Natl. Acad. Sci., 1990, 87, 682-685.Nitric oxide synthase converts ³ H-L-arginine into ³ H-L-citrullinewhich can be separated by cation exchange chromatography and quantifiedby scintillation counting.

Screen for neuronal nitric oxide synthase inhibiting activity

The enzyme is isolated from rat hippocampus or cerebellum. Thecerebellum or hippocampus of a male Sprague-Dawley rat (250-275 g) isremoved following CO₂ anaesthesia of the animal and decapitation.Cerebellar or hippocampal supernatant is prepared by homogenisation in50 mM Tris-HCl with 1 mM EDTA buffer (pH 7.2 at 25° C.) andcentifugation for 15 minutes at 20,000 g. Residual L-arginine is removedfrom the supernatant by chromatography through Dowex AG-50W-X8 sodiumform and hydrogen form columns successively, and further centrifugationat 1000 g for 30 seconds.

For the assay, 25 μl of the final supernatant is added to each of 96wells (of a 96 well filter plate) containing either 25 μl of an assaybuffer (50 mM HEPES, 1 mM EDTA, 1.5 mM CaCl₂, pH 7.4) or 25 μl of testcompound in the buffer at 22° C. and 25 μl of complete assay buffer (50mM HEPES, 1 mM EDTA, 1.5 mM CaCl₂, 1 mM DTT, 100 μM NADPH, 10 μg/mlcalmodulin, pH 7.4). Following a 10 minute equilibration period, 25 μlof an L-arginine solution (of concentration 18 μM ¹ H-L-arginine, 96 nM³ H-L-arginine) is added to each well to initiate the reaction. Thereaction is stopped after 10 minutes by addition of 200 μl of a slurryof termination buffer (20 mM HEPES, 2 mM EDTA, pH 5.5) and DowexAG-50W-X8 200-400 mesh.

Labelled L-citrulline is separated from labelled L-arginine by filteringeach filter plate and 75 μl of each terminated reaction is added to 3 mlof scintillation cocktail. The L-citrulline is then quantified byscintillation counting.

In a typical experiment using the cerebellar supernatant, basal activityis increased by 20,000 dpm/ml of sample above a reagent blank which hasan activity of 7,000 dpm/ml. A reference standard, N-nitro-L-arginine,which gives 80% inhibition of nitric oxide synthase at a concentrationof 1 μM, is tested in the assay to verify the procedure.

Screen for endothelial nitric oxide synthase inhibiting activity

The enzyme is isolated from human umbilical vein endothelial cells(HUVECs) by a procedure based on that of Pollock et al in Proc. Natl.Acad. Sci., 1991, 88, 10480-10484. HUVECs were purchased from CloneticsCorp (San Diego, Calif., USA) and cultured to confluency. Cells can bemaintained to passage 35-40 without significant loss of yield of nitricoxide synthase. When cells reach confluency, they are resuspended inDulbecco's phosphate buffered saline, centrifuged at 800 rpm for 10minutes, and the cell pellet is then homogenised in ice-cold 50 mMTris-HCl, 1 mM EDTA, 10% glycerol, 1 mM phenylmethylsulphonylfluoride, 2μM leupeptin at pH 4.2. Following centrifugation at 34,000 rpm for 60minutes, the pellet is solubilised in the homogenisation buffer whichalso contains 20 mM CHAPS. After a 30 minute incubation on ice, thesuspension is centrifuged at 34,000 rpm for 30 minutes. The resultingsupernatant is stored at -80° C. until use.

For the assay, 25 μl of the final supernatant is added to each of 12test tubes containing 25 μl L-arginine solution (of concentration 12 μM¹ H-L-arginine, 64 nM ³ H-L-arginine) and either 25 μl of an assaybuffer (50 mM HEPES, 1 mM EDTA, 1.5 mM CaCl₂, pH 7.4) or 25 μl of testcompound in the buffer at 22° C. To each test tube was added 25 μl ofcomplete assay buffer (50 mM HEPES, 1 mM EDTA, 1.5 mM CaCl₂, 1 mM DTT,100 μM NADPH, 10 μg/ml calmodulin, 12 μM tetrahydrobiopterin, pH 7.4) toinitiate the reaction and the reaction is stopped after 10 minutes byaddition of 2 ml of a termination buffer (20 mM HEPES, 2 mM EDTA, pH5.5).

Labelled L-citrulline is separated from labelled L-arginine bychromatography over a Dowex AG-50W-X8 200-400 mesh column. A 1 mlportion of each terminated reaction mixture is added to an individual 1ml column and the eluant combined with that from two 1 ml distilledwater washes and 16 ml of scintillation cocktail. The L-citrulline isthen quantified by scintillation counting.

In a typical experiment, basal activity is increased by 5,000 dpm/ml ofsample above a reagent blank which has an activity of 1500 dpm/ml. Areference standard, N-nitro-L-arginine, which gives 70-90% inhibition ofnitric oxide synthetase at a concentration of 1 μM, is tested in theassay to verify the procedure.

In the screens for nitric oxide synthase inhibition activity, compoundactivity is expressed as IC₅₀ (the concentration of drug substance whichgives 50% enzyme inhibition in the assay). IC₅₀ values for testcompounds were initially estimated from the inhibiting activity of 1, 10and 100 μM solutions of the compounds. Compounds that inhibited theenzyme by at least 50% at 10 μM were re-tested using more appropriateconcentrations so that an IC₅₀ could be determined.

When tested in the above screens, the compounds of Examples 1 to 6 belowshow IC₅₀ values for inhibition of neuronal nitric oxide synthase ofless than 10 μM and good selectivity for inhibition of the neuronalisoform of the enzyme, indicating that they are predicted to showparticularly useful therapeutic activity.

When compared with other compounds, the compounds of formula (I), andoptical isomers and racemates thereof and pharmaceutically acceptablesalts thereof, have the advantage that they may be less toxic, be moreefficacious, be longer acting, have a broader range of activity, be morepotent, be more selective for the neuronal isoform of nitric oxidesynthase enzyme, produce fewer side effects, be more easily absorbed orhave other useful pharmacological properties.

The invention is illustrated by the following examples:

EXAMPLE 1

N-(2-Methyl-1,2,3,4-tetrahydroisoguinolin-7-yl)-2-thiophenecarboximidamidedihydrochloride

a) 2-Methyl-7-nitro-1,2,3,4-tetrahydroisoguinoline hydrochloride

7-Nitro-1,2,3,4-tetrahydroisoquinoline (20 g, 93.2 mmol), formaldehyde(37% solution in water, 50 ml) and formic acid (90 ml) were heated atreflux for 1 h, cooled and poured over ice. The reaction mixture wasbasified with conc. ammonium hydroxide. The precipitated solid wascollected, dissolved in warm ethanol (200 ml), acidified with a mixtureof 95% ethanol-conc. HCl and the product was left to crystallize. Thetitle compound was obtained as a white solid (18.71 g, 87.8%), m.p.256-257° C.

b) 2-Methyl-1,2,3,4-tetrahydroisoguinolin-7-ylamine hydrochloride

2-Methyl-7-nitro-1,2,3,4-tetrahydroisoquinoline hydrochloride wasdissolved in methanol and hydrogenated at 50 psi in the presence of acatalytic quantity of 10% Pd-C. After 1 h the mixture was filteredthrough glass and evaporated to provide2-methyl-1,2,3,4-tetrahydroisoquinolin-7-ylamine hydrochloride, m.p.137-138° C.

c) 2-Thiophenecarboximidothioic acid, ethyl ester, hydrochloride

To a stirred solution of ethanethiol (28.4 g, 0.45 mols) in methylenechloride (500 ml) at 10° C. under nitrogen was added2-thiophenecarbonitrile (50.0 g, 0.45 mols). This solution was treatedwith a slow stream of HCl gas for 6 h. The reaction mixture was thenallowed to warm to room temperature for 18 h. Ether (200 ml) was addedand a white solid crystallized. The solid 2-thiophenecarboximidothioicacid, ethyl ester, hydrochloride was collected by filtration and airdried (65.8 g), m.p. 196-197° C.

d)N-(2-Methyl-1,2,3,4-tetrahydroisoguinolin-7-yl)-2-thiophenecarboximidamidedihydrochloride

2-Methyl-1,2,3,4-tetrahydroisoquinolin-7-ylamine hydrochloride (34.66 g)in 95% ethanol (600 ml) was warmed to 65° C. to dissolve most of thesolids, and the mixture was then allowed to cool with stirring. The nextday, the fine suspension of solids was treated with2-thiophenecarboximidothioic acid, ethyl ester, hydrochloride (41 g) andstirred at 23° C. All solids had dissolved by 2 h, and by 4 h new solidshad precipitated. The mixture was treated with concentrated hydrochloricacid (2 ml). The mixture was cooled to 0° C. and stirred for 30 minutes.The solids were filtered off, washed with ethanol (2×50 ml), and airdried to provideN-(2-methyl-1,2,3,4-tetrahydroisoquinolin-7-yl)-2-thiophenecarboximidamidedihydrochloride, m.p. 142-146° C.; MS ^(m) /z 272 [M+H]⁺.

EXAMPLE 2

N-(2-Isopropyl-1,2,3,4-tetrahydroisoquinolin-7-yl)-2-thiophenecarboximidamide

To a stirred solution ofN-(1,2,3,4-tetrahydroisoquinolin-7-yl)-2-thiophenecarboximidamide (7.0g, 21 mmol) in dimethylformamide (100 ml) was added potassium carbonate(14.6 g, 100 mmol). To this mixture was added 2-bromopropane (5.1 g, 42mmol), and the mixture was then heated to 40° C. for 72 h. The reactionmixture was poured into water (500 ml) and extracted with ethyl acetate(3×100 ml). The combined ethyl acetate extracts were washed with water(200 ml) and dried over magnesium sulfate. Evaporation of the solventyielded a crude oil, which was then dissolved in hot cyclohexane (250ml) and ethyl acetate (10 ml). Upon standing the title compoundcrystallized out and was collected by filtration (3.2 g), m.p. 110-111°C.

EXAMPLE 3

N-(2-Ethyl-1,2,3,4-tetrahydroisoguinolin-7-yl)-2-thiophenecarboximidamidehydrochloride

a) 2-Ethyl-7-nitro-1,2,3,4-tetrahydroisoguinoline hydrochloride

To 7-nitro-1,2,3,4-tetrahydroisoquinoline (5 g, 30 mmol) in acetonitrile(100 ml) was added ethyl methanesulfonate (6.38 g, 60 mmol) andpotassium carbonate (5 g). The mixture was heated to 40° C. for 18 h.The mixture was filtered and concentrated to an oil. The oil wasdissolved in methanol and treated with isopropanol-HCl. Thehydrochloride salt was collected by filtration (4.89 g, 67%), m.p.259-260° C.

b) 2-Ethyl-1,2,3,4-tetrahydroisoguinolin-7-ylamine hydrochloride

2-Ethyl-7-nitro-1,2,3,4-tetrahydroisoquinoline hydrochloride (4.89 g)was dissolved in methanol (250 ml) and hydrogenated at 50 psi in thepresence of a catalytic quantity of 5% Pd-C. After 1 h the mixture wasfiltered through glass and evaporated to an oil which was usedimmediately in the next step.

c) N-(2-Ethyl-1,2,3,4-tetrahydroisoguinolin-7-yl)2-thiophenecarboximidamide hydrochloride

To 2-ethyl-1,2,3,4-tetrahydroisoquinolin-7-ylamine hydrochloride (2.48g, 10 mmol) in isopropanol (25 ml) was added2-thiophenecarboximidothioic acid, methyl ester, hydroiodide (5.68 g, 20mmol). The mixture was heated to 50° C. for 24 h. The mixture was pouredinto water (50 ml), then basic water (150 ml). The mixture was extractedwith ethyl acetate (3×100 ml). The extracts were washed with water,dried with magnesium sulfate, filtered, and concentrated to an oil whichcrystallized upon standing. The solids were dissolved in ether andtreated with isopropanol-HCl. The solids were collected by filtration(1.41 g, 49%), m.p. 122-126° C.

EXAMPLE 4

N-(2-Propyl-1,2,3,4-tetrahydroisoquinolin-7-yl)-2-thiophenecarboximidamidedihydrochloride

a) 7-Nitro-2-propyl-1,2,3,4-tetrahydroisoguinoline hydrochloride

The title compound was prepared from7-nitro-1,2,3,4-tetrahydroisoquinoline (5 g, 30 mmol) and 1-bromopropane(7.36 g, 60 mmol) by a procedure analogous to that of Example 3(a). Thisyielded the hydrochloride salt (3.29 g, 43%), MS ^(m) /z 221 [M+H]⁺.

b) 2-Propyl-1,2,3,4-tetrahydroisoguinolin-7-ylamine hydrochloride

7-Nitro-2-propyl-1,2,3,4-tetrahydroisoquinoline hydrochloride (Example4(a), 3.29 g, 13 mmol) was hydrogenated using the process described inExample 3(b). The title compound hydrochloride salt thus obtained (3.07g, 100%) was used immediately in the next step.

c)N-(2-Propyl-1,2,3,4-tetrahydroisoguinolin-7-yl)-2-thiophenecarboximidamidedihydrochloride

2-Propyl-1,2,3,4-tetrahydroisoquinoline-7-ylamine hydrochloride (3.07 g,13 mmol) in DMF (30 ml) was treated with 2-thiophenecarboximidothioicacid, methyl ester, hydroiodide (2.84 g, 10 mmol) according to themethod of Example 3(c). The solids were recrystallized from ether (1.28g, 43%), MS ^(m) /z 300 [M+H]⁺. The dihydrochloride salt was made bydissolving these solids in ethanol, treating with ethanol-HCl, andtriturating with ethyl acetate (0.86 g, 73%), m.p. 241-243° C.

EXAMPLE 5

N-(2-Methyl-1,2,3,4-tetrahydroisoguinolin-7-yl)-3-thiophenecarboximidamidedihydrochloride

a) 3-Thiophenecarboximidothioic acid, methyl ester, hydroiodide

The title compound was prepared from 3-thiophenecarbothioamide andmethyl iodide by a procedure analogous to that described in Example 1(d)of WO 95/05363.

b)N-(2-Methyl-1,2,3,4-tetrahydroisoguinolin-7-yl)-3-thiophenecarboximidamidedihydrochloride

A mixture of 2-methyl-1,2,3,4-tetrahydroisoquinolin-7-ylaminehydrochloride (1.5 g, 7.55 mmol) and 3-thiophenecarboximidothioic acid,methyl ester, hydroiodide (2.69 g, 9.44 mmol) inN-methyl-2-pyrrolidinone (10 ml) was heated at 50° C. for 5 h. Theresulting solid mass was treated with isopropanol (50 ml), dissolved inwater, basified with conc. ammonium hydroxide and extracted twice withchloroform. The combined extracts were dried over magnesium sulphate,the solvent was evaporated and the residue was treated with ethanol-HClto give the title compound (1.37 g, 52%), MS ^(m) /z 272 [M+H]⁺.

EXAMPLE 6

N-(2-Butyl-1,2,3,4-tetrahydroisoguinolin-7-yl)-2-thiophenecarboximidamide

N-(1,2,3 ,4-Tetrahydroisoquinolin-7-yl)-2-thiophenecarboximidamide (3.0g, 9 mmol) and 1 -chlorobutane (1.67 g, 18 mmol) were reacted togetheraccording to the method of Example 2 except that 95% ethanol was used asthe solvent. The crude oil thus obtained was chromatographed on silicagel eluting with 10% methanol-chloroform to give an oil which wascrystallized from hot hexane (1.12 g, 40%), m.p. 95-96° C.

What is claimed is:
 1. A compound of formula (I) ##STR13## wherein: R¹represents a 2-thienyl or 3-thienyl ring; andR² represents C₁₋₄ alkyl;or an optical isomer or racemate thereof or a pharmaceuticallyacceptable salt thereof.
 2. A compound of formula (I), according toclaim 1, wherein R¹ represents 2-thienyl.
 3. A compound of formula (I),according to claim 1, wherein R² represents methyl.
 4. A compound offormula (I), according to claim 1, wherein R¹ represents 2-thienyl andR³ represents methyl.
 5. A compound of formula (I) whichis:N-(2-methyl-1,2,3,4-tetrahydroisoquinolin-7-yl)-2-thiophenecarboximidamide;N-(2-isopropyl-1,2,3,4-tetrahydroisoquinolin-7-yl)-2-thiophenecarboximidamide;N-(2-ethyl-1,2,3,4-tetrahydroisoquinolin-7-yl)-2-thiophenecarboximidamide;N-(2-propyl-1,2,3,4-tetrahydroisoquinolin-7-yl)-2-thiophenecarboximidamide;N-(2-methyl-1,2,3,4-tetrahydroisoquinolin-7-yl)-3-thiophenecarboximidamide;N-(2-butyl-1,2,3,4-tetrahydroisoquinolin-7-yl)-2-thiophenecarboximidamide;oran optical isomer or racemate of any one thereof or a pharmaceuticallyacceptable salt of any one thereof.
 6. A pharmaceutical formulationcomprising a compound of formula (I), as defined in claim 1, or anoptical isomer or racemate thereof or a pharmaceutically acceptable saltthereof, optionally in admixture with a pharmaceutically acceptablediluent or carrier.
 7. A pharmaceutical formulation comprising acompound of formula (I), as defined in claim 1, or an optical isomer orracemate thereof or a pharmaceutically acceptable salt thereof, incombination with L-Dopa, or with an opiate analgesic, optionally inadmixture with a pharmaceutically acceptable diluent or carrier.
 8. Amethod of treating, or reducing the risk of, a human disease orcondition in which inhibition of nitric oxide synthase activity isbeneficial which comprises administering to a person suffering from orsusceptible to such a disease or condition, a therapeutically effectiveamount of a compound of formula (I), as defined in claim 1, or anoptical isomer or racemate thereof or a pharmaceutically acceptable saltthereof.
 9. A method of treatment according to claim 8 in which it ispredominantly the neuronal isoform of nitric oxide synthase that isinhibited.
 10. A method of treating, or reducing the risk of, hypoxia orstroke or ischaemia or neurodegenerative conditions or schizophrenia orpain or migraine, or for the prevention and reversal of tolerance toopiates and diazepines, or for the treatment of drug addiction whichcomprises administering to a person suffering from or susceptible tosuch a disease or condition a therapeutically effective amount of acompound of formula (I), as defined in claim 1, or an optical isomer orracemate thereof or a pharmaceutically acceptable salt thereof.
 11. Amethod of treatment according to claim 10, wherein the condition to betreated is selected from the group consisting of hypoxia, ischaemia,stroke, Huntington's disease, Parkinson's disease, Amyotrophic LateralSclerosis, schizophrenia and pain.
 12. A method of treatment accordingto claim 11, wherein the condition to be treated is stroke.
 13. A methodof treatment according to claim 11, wherein the condition to be treatedis Amyotrophic Lateral Sclerosis.
 14. A method of treatment according toclaim 11, wherein the condition to be treated is pain.
 15. A method oftreatment according to claim 11, wherein the condition to be treated isHuntington's disease.
 16. A method of treatment according to claim 11,wherein the condition to be treated is Parkinson's disease.
 17. A methodof treatment according to claim 11, wherein the condition to be treatedis schizophrenia.
 18. A method of treating, or reducing the risk ofsuffering from, pain which comprises administering to a person sufferingfrom or at risk of suffering from, pain a therapeutically effectiveamount of a compound of formula (I), as defined in claim 1, or anoptical isomer or racemate thereof or a pharmaceutically acceptable saltthereof, in combination with an opiate analgesic agent.
 19. A method oftreatment of Parkinson's disease which comprises administering to aperson suffering from, or at increased risk of suffering from,Parkinson's disease, a therapeutically effective amount of a compound offormula (I), as defined in claim 1, or an optical isomer or racematethereof or a pharmaceutically acceptable salt thereof, in combinationwith L-Dopa.
 20. A process for the preparation of a compound of formula(I), as defined in claim 1, and optical isomers and racemates thereofand pharmaceutically acceptable salts thereof, which comprises:(a)preparing a compound of formula (I) by reacting a corresponding compoundof formula (II) ##STR14## wherein R² is as defined in claim 1, with acompound of formula (III) or an acid addition salt thereof ##STR15##wherein R¹ is as defined in claim 1 and L is a leaving group; (b)preparing a compound of formula (I) by reacting a corresponding compoundof formula (IV) ##STR16## wherein R² is as defined in claim 1 and HA isan acid, with a compound of formula (V) ##STR17## wherein R¹ is asdefined in claim 1; (c) preparing a compound of formula (I) by reactinga compound of formula (VI) ##STR18## wherein R¹ is as defined in claim1, with a compound of formula (VII)

    R.sup.2 --L                                                (VII)

wherein R² is as defined in claim 1 and L is a leaving group; or (d)preparing a compound of formula (I) in which R² represents methyl byreacting a compound of formula (VI) with formaldehyde and formic acid;and where desired or necessary converting the resultant compound offormula (I), or another salt thereof, into a pharmaceutically acceptablesalt thereof, or vice versa, and where desired converting the resultantcompound of formula (I) into an optical isomer thereof.
 21. Apharmaceutical formulation according to claim 7, wherein said opiateanalgesic is morphine.
 22. A method of treatment according to claim 18,wherein said opiate analgesic agent is morphine.
 23. A method oftreatment according to claim 10, wherein the condition to be treated ismigraine.